A comprehensive analysis of CEBPA on prognosis and function in uterine corpus endometrial carcinoma

Abstract

Uterine corpus endometrial carcinoma (UCEC) is one of the most common tumours of the female reproductive system. CCAAT enhancer-binding protein alpha (CEBPA) is a member of the transcription factor family involved in regulating processes such as cell proliferation, differentiation, metabolism, and the immune response. However, the role of CEBPA in UCEC has not been clarified. Here, we performed a comprehensive analysis to explore the expression level, prognostic value, immune infiltration and biological function of CEBPA in UCEC. In this study, we found that CEBPA expression was upregulated and associated with poor prognosis in UCEC patients. KEGG and GO analyses revealed that the genes positively correlated with CEBPA were enriched primarily in immune regulation and oxidative phosphorylation. Immune infiltration analysis revealed that CEBPA is strongly correlated with immune cell infiltration in UCEC. RT-qPCR indicated that CEBPA may regulate the OXPHOS level in Ishikawa cells. CCK-8, cell cycle, Transwell and scratch wound healing assays revealed that CEBPA promoted Ishikawa cell proliferation, invasion and migration. In addition, PPI and survival analyses suggested that CEBPG may be a potential target of CEBPA in UCEC. These results demonstrated that CEBPA may be a potential therapeutic target in UCEC.

Keywords: CEBPA; Immune filtration; Invasion; Migration; Proliferation; UCEC.

Fig. 1

Expression level and prognostic value of CEBPA in UCEC. (a–b) CEBPA mRNA expression levels in UCEC tumour tissues and normal tissues from TCGA and GTEx. (c) CEBPA protein expression levels in UCEC tumour tissues and normal endometrial tissues from the HPA website (× 200) (immunohistochemistry, hematoxylin-eosin staining). (d) Prognostic significance of CEBPA expression in UCEC according to Kaplan–Meier plotter curves. OS, overall survival. (e) The predictive value of CEBPA for the prognosis of UCEC according to ROC curve. *P < 0.05, **P < 0.01, ***P < 0.001.

Fig. 2

KEGG and GO enrichment of genes related to the expression of CEBPA in UCEC. (a) Heatmap showing the enrichment patterns of the top 50 genes positively correlated with the CBEPA. (b) Heatmap showing the enrichment patterns of the top 50 genes negatively correlated with the CBEPA. (c) GO-BP enrichment analysis was applied to the top 100 genes that were positively correlated with the CBEPA. (d) KEGG enrichment analysis was applied to the top 100 genes that were positively correlated with the CBEPA.

Fig. 3

Relationship between CEBPA expression and immune cell infiltration in UCEC. (a) Relative proportions of immune cell infiltration between the high CEBPA expression group and the low CEBPA expression group in UCEC. (b) Diagram of the difference in immune cell infiltration proportions between the high CEBPA expression group and the low CEBPA expression group. (c) Correlation heatmap among immune cell populations; red and blue indicate positive and negative correlations, respectively. (d) Lollipop chart shows the relationships between CEBPA and immune cell infiltration. (e–h) M1 macrophages, M2 macrophages, memory B cells, and gamma delta T cells. (i–k) naive B cells, resting memory CD4 T cells, and activated mast cells.

Fig. 4

Effects of CEBPA on the oxidative phosphorylation level in Ishikawa cells. (a–b) The efficiency of CEBPA overexpression in Ishikawa cells was determined via RT-qPCR and western blotting. (c–d) The efficiency of CEBPA knockdown in Ishikawa cells was determined via RT-qPCR and western blotting. (e) Expression of the core members of OXPHOS in OE-CEBPA Ishikawa cells and control cells via RT-qPCR. (f) Expression of the core members of OXPHOS in sh-CEBPA Ishikawa cells and control cells via RT-qPCR. *P < 0.05, **P < 0.01, ***P < 0.001.

Fig. 5

Effects of CEBPA on the proliferation, invasion and migration ability of Ishikawa cells. (a–d) CCK-8 and cell cycle assays were performed to evaluate the proliferation of sh-CEBPA Ishikawa cells, OE-CEBPA Ishikawa cells, and control cells. (e–f) Scratch wound healing assay was applied to evaluate the migration of sh-CEBPA Ishikawa cells, OE-CEBPA Ishikawa cells, and control cells. (g–h) Transwell assay was performed to determine the invasiveness of sh-CEBPA Ishikawa cells, OE-CEBPA Ishikawa cells, and control cells. *P < 0.05, **P < 0.01, ***P < 0.001.

Fig. 6

Screening and validation of CEBPA target genes in UCEC. (a–b) The intersection between genes that were positively or negatively related to CEBPA, hTF target genes and footprintDB target genes. (c) The PPI network map of 42 intersecting genes in the STRING portal. (d) Comparison of CEBPA-related genes expression between normal tissue and tumour tissue in the TCGA-UCEC cluster. (e) The impact of APOE, CEBPG and SPI1 on the overall survival (OS) of UCEC patients was determined via Kaplan–Meier plotter curves. (f) CEBPG mRNA expression levels in UCEC tumour tissues and normal tissues from TCGA and GTEx. (g) mRNA expression of CEBPG in Ishikawa cells after silencing or overexpressing CEBPA via RT-qPCR. *P < 0.05, **P < 0.01, ***P < 0.001.